The following abbreviations are used in this disclosure: CAM, cell adhesion molecules; EBM, endothelial basal medium; EC, endothelial cells; EDTA, ethylene-diaminetetraacetic acid; EGF, epidermal growth factor; ELAM-1, endothelial leukocyte adhesion molecule-1; FBS, fetal bovine serum; HEV, high endothelial venules; HUVEC, human umbilical vein endothelial cells; ICAM-1, intracellular adhesion molecule-1; IL-1, interleukin-1; IL-1β, interleukin-1-beta; IL-4, interleukin-4; INF-γ, interferon-gamma; LDL, low density lipoprotein; LFA-1, lymphocyte function-associated antigen-1; LTBMC, long-term bone marrow culture system; mAb, monoclonal antibody; MLN, mesenteric lymph node; PBL, peripheral blood lymphocytes; PBS, phosphate-buffered saline; PMN, polymorphonuclear leukocyte; SEM, standard deviation; TNF-α, tumor necrosis factor-alpha; VCAM-1, vascular cell adhesion molecule-1; VLA-4, leukocyte integrin VLA-4; WM, Waymouth medium. Throughout the specification, the notation “(#)” is used to refer to the documents listed in the appended Citations section.
Migration of lymphocytes from the bloodstream into surrounding tissues is a dynamic, multistep process initiated by attachment to the luminal surface of endothelial cells (EC) lining the postcapillary venules. Certain components of the microvasculature, notably the morphologically distinct high endothelial venules (HEV) found in lymphoid organs such as lymph nodes. Peyer's patches, and tonsils, continuously support lymphocyte binding and transmigration. Some adhesive interactions attendant with movement into these sites are, at least operationally, organ-specific (1-6). Others are mediated by cell adhesion molecules (CAM) that have more general tissue distributions, for example, ICAM-1/LFA-1 interactions (7-9). During both acute and chronic inflammation, microvascular endothelial cells at other sites can be induced to support traffic of various leukocyte subtypes (5, 10). Accumulation of lymphocytes in chronic inflammations, e.g., arthritic synovia, is usually accompanied by conversion of the local postcapillary venules to a cuboidal morphology and expression of new adhesive structures (3, 5). It has been suggested that lymphocyte adhesion to endothelial cells in chronic inflammatory lesions also incorporates an element of organ- or site-specificity (11). The complete identity and balance of inductive factors in the local microenvironment that contribute to the endothelial-cell “traffic” phenotype, and particularly its organ-specific character at some sites, have yet to be defined in molecular terms. Other factors are likely to be important, e.g., endothelial cell contact with the underlying extracellular matrix (12); but, clearly, release of proinflammatory cytokines in the local milieu contributes markedly to the upregulation of cell adhesion molecules on endothelial cells (13, 14).
For example, interleukin-1 (IL-1), TNF-α, and IFN-γ have all been shown to increase adhesiveness of cultured endothelial cells for granulocytes and lymphocytes (15-23). In some cases these effects are paralleled by enhanced leukocyte migration to sites of cytokine injection in vivo (24, 25). Recently, much progress has been made in identification of specific cell adhesion molecules induced on endothelial cells by proinflammatory cytokines. IL-1, for example, induces endothelial leukocyte adhesion molecule-1 (ELAM-1), a member of the LEC-CAM (Lectin, EGF, Complement-Cellular Adhesive Molecule) family (19, 26), which is selectively adhesive for polymorphonuclear leukocytes and weakly adhesive for monocytes. Similarly, cytokine induction of intercellular adhesion molecule-1 (ICAM-1), a ligand for the leukointegrin LFA-1 (lymphocyte function-associated antigen-1), has been reported on endothelial cells (27). Recently, vascular cell adhesion molecule-1 (VCAM-1) was identified as a TNF- and IL1-inducible ligand for VLA4-mediated attachment of lymphocyte adhesion to human umbilical vein endothelial cells (HUVEC) (28-30). Although not directly linked functionally to lymphocyte transmigration, other cell surface markers associated with traffic endothelium in vivo have been shown to be induced by IFN-γ (15). Additional adhesive ligands of more limited tissue distribution, termed vascular addressins, MECA-79 and MECA-367, have been identified in lymph nodes and mucosal lymphoid tissues, respectively (31, 32). Whether these ligands can be induced in vitro by specific cytokines is not known at this point, but studies of transgenic mice suggest that IFN-γ may contribute to their expression in vivo (33).
The capacity of cytokines to enhance lymphocyte adhesion to microvascular-derived endothelial cells has been analyzed in rodents and sheep (15, 24, 25). In humans, wherein most of the molecular definition of EC-CAM exists, cytokine induction has been studied almost exclusively using umbilical vein as the endothelial cell source (16-19). As pointed out recently by Issekutz (24), certain disparities exist between results obtained in these systems.
Because of this and since our preliminary results indicated that large-vessel- and microvascular-derived endothelial cells might differ in cytokine responses via-á-vis adhesive events, we endeavored to test how immunologically active cytokines affected lymphocyte adhesion to primate (macaque) microvascular endothelial cells. Further, since there have been suggestions of cytokine dependence for the traffic endothelial cell phenotype not only at sites of inflammation, but also for high endothelium in lymph nodes (34), mesenteric lymph nodes were used as one source of microvascular endothelial cells.